The long-term objectives of this proposal are to elucidate the functional cooperation between BRCA1 and a novel transcriptional corepressor in hormonal gene regulation. In contrast to the strides taken in studying the mechanisms of hormonal gene activation, the molecular basis for counter-balancing ligand-dependent gene expression remains to be elucidated. Recent studies indicate that BRCA1 interacts with ER-alpha and represses its function in gene activation, which could explain the tissue-specific effect of BRCA1 mutations. However, BRCA1 mutations only account for a small proportion of breast cancer. It is therefore possible that defects of other players in the BRCAl-regulated process may also contribute to breast cancer development. Cofactor of BRCA1 (COBRA1) was identified based upon its ability to interact with BRCAI. Like BRCA1, COBRA1 also directly binds to ER-alpha and inhibits the ligand-dependent gene activation. COBRA1 is selectively recruited to a subset of ER-responsive promoters in a ligand-dependent manner. As an integral subunit of a negative transcription elongation complex, COBRA1 mediates stalling of RNA polymerase II (RNAinvestigatorI). Preliminary work also indicates that COBRA1 inhibits the growth of breast epithelial cells. Based on the above-mentioned findings, it is hypothesized that COBRA1 may act in concert with BRCA1 to repress ER-mediated gene expression, and that loss of COBRA1 function may contribute to an elevated risk of breast cancer. The specific aims of the proposal are: 1) to determine the biochemical basis for the promoter-specific action of COBRA1. The impact of COBRA1 on ER-alpha binding to DNA and other known coregulators including BRCA1 will be assessed. 2) To identify the physiological targets for COBRA1 regulation. Primary efforts will be focused on the endogenous ER-responsive genes. Recruitment of COBRA1 to native promoters and its impact on the dynamics of transcription complexes will be analyzed. 3) To assess the possible cooperation between COBRA1 and BRCA1. Cell and molecular biological assays will be utilized to determine a potential synergy between COBRA1 and BRCA1 in gene regulation and growth control of mammary epithelial cells. These studies promise to provide important insight into hormonal gene regulation and suppression of breast cancer development.